Method and kit for determining quantity of protein

Abstract

A method for determining the quantity of a protein includes the steps of: (a) preparing a crude protein liquid containing crude proteins by solubilizing tissue or cells in a pretreatment liquid; (b) contacting the prepared crude protein liquid with a solid phase to allow the crude proteins to bind to the solid phase; and (c) determining the quantity of a target protein in the crude proteins binding to the solid phase using a substance which binds specifically to the target protein.

What is claimed is:  
     
         1 . A method for determining the quantity of a protein comprising the steps of: 
 (a) preparing a crude protein liquid containing crude proteins by solubilizing tissue or cells in a pretreatment liquid;    (b) contacting the prepared crude protein liquid with a solid phase to allow the crude proteins to bind to the solid phase; and    (c) determining the quantity of a target protein in the crude proteins binding to the solid phase by using a substance which binds specifically to the target protein.    
     
     
         2 . The method according to  claim 1 , wherein the pretreatment liquid contains a surfactant.  
     
     
         3 . The method according to  claim 2 , wherein a concentration of the surfactant in the pretreatment liquid is 1.0 to 0.01 w/v %.  
     
     
         4 . The method according to  claim 1 , wherein the pretreatment liquid contains a protease inhibitor.  
     
     
         5 . The method according to  claim 1 , wherein the solid phase comprises a porous membrane or beads.  
     
     
         6 . The method according to  claim 1 , wherein the substance which binds specifically to the target protein is an antibody or a nucleic acid.  
     
     
         7 . The method according to  claim 1 , wherein the target protein is a water-soluble protein.  
     
     
         8 . The method according to  claim 1 , wherein the target protein is at least one of actin, Cdk1, Cdk2, Cdk4, Cdk6, Cycline B, Cycline D, Cycline E, P16, P21, P27 and C-myc.  
     
     
         9 . The method according to  claim 1 , wherein the step (b) comprises the steps of putting the crude protein liquid in one or more wells of a plate provided with a porous membrane as the solid phase at the bottom of the wells and allowing the crude proteins to bind to the porous membrane by suction from a porous membrane side of the plate.  
     
     
         10 . The method according to  claim 9 , wherein the porous membrane is a hydrophobic porous membrane.  
     
     
         11 . The method according to  claim 9 , wherein the substance which binds specifically to the target protein is an antibody or a nucleic acid.  
     
     
         12 . The method according to  claim 9 , wherein the plate has two or more wells.  
     
     
         13 . The method according to  claim 12 , wherein the step (c) comprises the step of supplying the wells with the substance which binds specifically to the target protein, a first one of the wells is supplied with a substance which binds specifically to a first target protein, and a second one of the wells is supplied with a substance which binds specifically to a second target protein.  
     
     
         14 . The method according to  claim 13 , which is for determining the quantity of two or more proteins.  
     
     
         15 . A diagnostic method for diagnosing a disease, the method comprising the steps of: 
 (a) preparing a crude protein liquid containing crude proteins by solubilizing tissue or cells in a pretreatment liquid;    (b) contacting the prepared crude protein liquid with a solid phase to allow the crude proteins to bind to the solid phase; and    (c) determining the quantity of a target protein in the crude proteins binding to the solid phase by using a substance which binds specifically to the target protein; and    (d) diagnosing a disease according to the determined quantity of the target protein.    
     
     
         16 . The diagnostic method according to  claim 15 , wherein the step (b) comprises the steps of putting the crude protein liquid in one or more wells of a plate provided with a porous membrane as the solid phase at the bottom of the wells and allowing the crude proteins to bind to the porous membrane by suction from a porous membrane side of the plate.  
     
     
         17 . The diagnostic method according to  claim 15 , wherein the disease is stomach cancer, intestinal cancer, breast cancer, lung cancer, esophagus cancer, prostate cancer, liver cancer, kidney cancer, bladder cancer, skin cancer, uterine cancer, brain tumor, osteosarcoma or myeloma.  
     
     
         18 . The diagnostic method according to  claim 15 , wherein the target protein is at least one of actin, Cdk1, Cdk2, Cdk4, Cdk6, Cycline B, Cycline D, Cycline E, P16, P21, P27 and C-myc.  
     
     
         19 . A kit for determining the quality of a protein comprising: 
 a pretreatment liquid for solubilizing tissue or cells;    a solid phase for binding crude proteins in a crude protein liquid prepared by solubilizing the tissue or cells thereto; and    a substance which binds specifically to a target protein in the crude proteins.    
     
     
         20 . The kit according to  claim 19 , wherein the solid phase is a porous membrane or beads.